The ultimate goal of this proposal is to leverage Vstat120 expressing oncolytic HSV-1 viruses (oHSV) for glioma therapy. Vstat120 is the extracellular fragment of Brain Angiogenesis Inhibitor 1 (BAI1) that has potent anti-angiogenic and anti-tumor effects. During the last cycle we created two Vstat120 expressing oncolytic viruses in different virus backbones. RAMBO, the first generation Vstat120 expressing virus, was created in a doubly attenuated virus back bone that is similar to G207, which has been tested in patients and found to be safe. RAMBO showed significantly better anti-tumor efficacy in mice with established brain tumors compared to the control virus with an identical backbone but lacking Vstat120 expression [5]. 34.5ENVE, the second generation Vstat120 expressing virus, was created in a virus backbone that is transcriptionally driven under the control of a nestin promoter. 34.5ENVE showed the best efficacy in GBM models that expressed high nestin levels. Given the significant improvement in anti-tumor efficacy of 34.5ENVE we have pursued its translational development with NIH (NCI NeXT program and NINDS CREATE program). The concern articulated by advisors at both NINDS and NCI was the fact that nestin is expressed in some normal cells, prompting us to reconsider tighter of nestin driven ICP34.5 expression. Here we propose to (Aim 1) modulate the backbone of 34.5ENVE to precisely control its replication in tumor cells and minimize toxicity to normal brain neurons and neural stem cells. We will further (Aim 2) evaluate the immunological consequences of this virus alone and (Aim 3) in conjunction with proteasome inhibition. At the conclusion of this project we will have an optimized oncolytic HSV vector that expresses Vstat120 which can be pursued for translational development with NIH.